Multiple drug resistant tuberculosis (MDRTB) has recently emerged as a serious public health concern. To develop better drugs or immunotherapies to treat MDRTB and design improved diagnostic protocols, it is imperative that drug targets and drug resistance mechanisms of M. tuberculosis are defined. Our laboratory has investigated the mechanism of resistance to Isoniazid, rifampin and streptomycin in 53 clinical M. tuberculosis isolates. We have demonstrated that reduced sensitivity to INH often is associated with alterations in the catalase-peroxidase (kat G) locus or in the promoter of the inh A gene. Resistance to rifampin is conferred by mutations in the rpo B gene (RNA polymerase). Genetic alterations in the rps L (ribosomal S12 protein) on the 16S rRNA genes are primarily responsible for streptomycin resistance in MDR TB strains. Furthermore, we established that multiple drug resistance in M. tuberculosis results from an accumulation of mutations in genes encoding drug targets and not from a novel pleiotropic resistance determinant. A recent focus of our research has been the molecular characterization of the catalase puoxidine (katG) protein that has been shown to be associated with mycobacterium virulence as well as sensitivity to INH using site-directed mutagenesis. We are currently examining the effects of specific katG mutations on the biological role of the protein as a virulence factor and a drug resistance modulator.